The overall goal of this Phase I SBIR application is to develop a non-cell-based means of inhibiting coagulation and platelet activation on engineered vascular grafts, so that small-diameter arterial grafts may be available off the shelf. This arterial graft will consist of human engineered tissue that is decellularized and coated with a novel, covalently-bound layer that mimics aspects of the endothelial cell glycocalyx. By providing an anti-thrombotic function through inhibition of the intrinsic clotting pathway and platelets, this coating will address the main problem with current small diameter vascular grafts, which is early, severe clotting. If functional and successful, this engineered, coated arterial graft will provide an option for patients who lack available vein for bypass conduit, and possibly eliminate the need to harvest a patient's own vein tissue for use as a replacement conduit in vascular bypass surgeries. Cardiovascular disease is the most costly and deadly disease in the US and in much of the Western world. Peripheral vascular disease (PAD) affects 12-20% of people over age 65. Symptomatic but stable disease is usually treated with conservative therapies such as dietary changes, exercise and pharmacological therapies. However, 20-30% of PAD patients suffer from debilitating claudication or critical limb ischemia, and in these patients, limb revascularization offers relief of symptoms and prevention of amputation. Catheter- based therapies are being used to restore arterial patency in PAD patients, but these techniques often offer only temporary relief, and many patients eventually require surgery. Currently, there are roughly 130 peripheral arterial bypass procedures performed annually for every 100,000 Medicare beneficiaries, corresponding to approximately 61,000 peripheral bypass operations each year in the US. Complicating this situation, almost one-third of patients requiring bypass do not possess suitable autologous vein for use in their bypass procedure due to prior vein harvest, venous disease, or the desire to preserve vein for anticipated coronary or other bypass procedures. Synthetic grafts composted of PTFE have been used in place of native vein, but the patency of PTFE grafts in the peripheral circulation is only 50-60% at 3 years. Heparin-coating of PTFE grafts in order to attempt to reduce failure through thrombosis, as in the Gore Propaten(R) graft, has not resulted in substantial improvement, and these heparin coated PTFE grafts still display lower patency rates than autologous vein. Hence, there is a significant medical need for a small-diameter, off the shelf arterial graft that eliminates the need for vein harvest yet functions better than PTFE, and ideally as well as native vein, in the peripheral circulation without clotting. Such a product could substantially improve outcomes for patients with severe PAD. Humacyte has developed methods to grow engineered vascular tissues from banked human smooth muscle cells that are seeded onto a biodegradable scaffold and cultured in bioreactors. No cells are harvested from the recipient for this process. These tissues are then decellularized, creating an acellular tubular tissue that has excellent mechanical characteristics. We have tested these tubular engineered tissues as arteriovenous grafts in a primate model, and they have shown excellent function. Clinical trials of the Humacyte graft as an arteriovenous conduit for hemodialysis access are currently underway in the United States and Europe. In this Phase I SBIR application, this novel graft will be further enhanced with a creative new, covalently-bound luminal coating that mimics aspects of the endothelial glycocalyx, thereby increasing the graft's thromboresistance and allowing the graft to remain patent even when used in small-diameter settings, such as bypass procedures for PAD. In this application, we will test the ability of the coating to withstand the shear stress present in the peripheral circulation and test the stability and functionality of this coating after a period of storage in saline in order to ascertain whether the coating has an acceptable shelf-life. Finally, we will determine the in vivo efficacy of coated grafts in a short-term (2-week) porcine carotid artery bypass model. If this coating strategy is successful, then the chemistry developed herein could not only create the first graft to function well in small-diameter PAD bypass, but it could pave the way for using the Humacyte HAVG in coronary artery bypass. In addition, the chemistry could be suitably modified to covalently coat other blood-contacting devices, thereby decreasing thrombogenicity and improving outcomes for a variety of cardiovascular implants and devices. Thus, this novel approach could have a significant impact on the field of vascular surgery.